Publications

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    ABSTRACT: Background. NOS/(•)NO inhibitors are potential therapeutics for sepsis, yet they increase clinical mortality. However, there has been no in vivo investigation of the (in vitro) (•)NO scavenger, cobalamin's (Cbl) endogenous effects on NOS/(•)NO/inflammatory mediators during the immune response to sepsis. Methods. We used quantitative polymerase chain reaction (qPCR), ELISA, Western blot, and NOS Griess assays, in a C57BL/6 mouse, acute endotoxaemia model. Results. During the immune response, pro-inflammatory phase, parenteral hydroxocobalamin (HOCbl) treatment partially inhibits hepatic, but not lung, iNOS mRNA and promotes lung eNOS mRNA, but attenuates the LPS hepatic rise in eNOS mRNA, whilst paradoxically promoting high iNOS/eNOS protein translation, but relatively moderate (•)NO production. HOCbl/NOS/(•)NO regulation is reciprocally associated with lower 4 h expression of TNF-α, IL-1 β , COX-2, and lower circulating TNF-α, but not IL-6. In resolution, 24 h after LPS, HOCbl completely abrogates a major late mediator of sepsis mortality, high mobility group box 1 (HMGB1) mRNA, inhibits iNOS mRNA, and attenuates LPS-induced hepatic inhibition of eNOS mRNA, whilst showing increased, but still moderate, NOS activity, relative to LPS only. experiments (LPS+D-Galactosamine) HOCbl afforded significant, dose-dependent protection in mice Conclusions. HOCbl produces a complex, time- and organ-dependent, selective regulation of NOS/(•)NO during endotoxaemia, corollary regulation of downstream inflammatory mediators, and increased survival. This merits clinical evaluation.
    Mediators of Inflammation 01/2013; 2013:741804. · 3.88 Impact Factor
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    Carmen Wheatley
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    ABSTRACT: Mammalian nitric oxide synthases (NOS) are a source of the universal second messenger, and pivotal biochemical molecule, nitric oxide (. NO). NOS are assumed to function catalytically in a haem-centred manner, by analogy with cytochrome P450. Yet, they differ significantly. Cobalamin, vitamin B12, is believed to function almost solely as an . NO scavenger and, latterly, as a direct, physiological inhibitor of the NOS. Yet, in pathology, associated to cobalamin deficiency, functional or otherwise, NOS over-produce superoxide, peroxynitrite (ONOO -), and other reactive nitrite species, rather than . NO (Figure 7). This paper offers a radical, new solution to the gaps and inconsistencies in the current understanding of the mechanism of haem-centred NOS catalysis, which also challenges the other existing paradigm of cobalamin as just an . NO mop. Examination of a wide diversity of NOS and cobalamin-dependent enzyme structure-function studies, as well as data from the . NO/cobalamin chemical, biochemical, immunological, genetic, and clinical literature, offers indications that cobalamin, specifically, in one of its active forms, adenosylcobalamin (AdoCbl), may have a third, eukaryotic coenzyme function as the principal cofactor of well-regulated NOS catalysis in vivo. The AdoCbl-centred NOS reaction is described in detail (Figure 5), and some existing evidence that, in vitro, without AdoCbl, NOS turnover activity is significantly slower than in in vivo AdoCbl-rich environments, is presented. AdoCbl, in conjunction with tetrahydrobiopterin, couples NOS oxygen binding/activation to L-arginine hydroxylation and . NO synthesis much more effectively than does haem, overcoming NOS spatial and redox problems, leading to productive catalysis, decreased radical formation/escape, with a consequent increased ratio of . NO to ONOO -, and prevention of pathology (Figures 5 & 7). In vivo, haem-centred NOS catalysis may, in fact, be the back-up NOS reaction, and it"s predominance in the absence of AdoCbl, with a consequent lowering of the . NO/ONOO-ratio, is the real source of supposedly . NO derived pathology.
    Hypotheses in the Life Sciences. 10/2012; 2(2-2042 8960):31-54.
  • Carmen WHEATLEY, Mauro PERRETTI, Fulvio D'ACQUISTO
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    ABSTRACT: The present invention provides methods of treatment and uses of thiolatocobalamin compounds for diseases characterised by NOS depression, malfunction, or dysregulation, for example inflammation, the inflammatory phase of the immune response, SIRS/sepsis/septic, traumatic, anaphylactic shock, a bacterial, viral or parasitidal disease, cancer, spinal injury or cardiovascular disease.
    Ref. No: WO/2009/071905, Year: 06/2009
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    Carmen Wheatley
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    ABSTRACT: Several mysteries surround the structure and function of the nitric oxide synthases (NOS). The NOS oxygenase domain structure is unusually open with a large area of solvent that could accommodate an unidentified ligand. The exact mechanism of the two-step five-electron monoxygenation of arginine to N(G)-hydroxy-L-arginine, thence to citrulline and nitric oxide (NO), is not clear, particularly as arginine/N(G)-hydroxy-L-arginine is bound at a great distance to the supposed catalytic heme Fe [III], as the anti-stereoisomer. The Return of the Scarlet Pimpernel Paper proposed that cobalamin is a primary indirect regulator of the NOS. An additional direct regulatory effect of the 'base-off' dimethylbenzimidazole of glutathionylcobalamin (GSCbl), which may act as a sixth ligand to the heme iron, promote Co-oriented, BH(4)/BH(3) radical catalysed oxidation of L-arginine to NO, and possibly regulate the rate of inducible NOS/NO production by the NOS dimers, is further advanced. The absence of homology between the NOS and methionine synthase/methylmalonyl CoA mutase may enable GSCbl to regulate both sets of enzymes simultaneously by completely separate mechanisms. Thus, cobalamin may exert central control over both pro-and anti-inflammatory systems.
    Journal of Nutritional & Environmental Medicine 02/2007; 16(3-4):212-226.
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    Carmen Wheatley
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    ABSTRACT: The up-regulation of transcobalamins [hitherto posited as indicating a central need for cobalamin (Cbl) in inflammation], whose expression, like inducible nitric oxide synthase (iNOS), is Sp1- and interferondependent, together with increased intracellular formation of glutathionylcobalamin (GSCbl), adenosylcobalamin (AdoCbl), methylcobalamin (MeCbl), may be essential for the timely promotion and later selective inhibition of iNOS and concordant regulation of endothelial and neuronal NOS (eNOS/nNOS.) Cbl may ensure controlled high output of nitric oxide (NO) and its safe deployment, because: (1) Cbl is ultimately responsible for the synthesis or availability of the NOS substrates and cofactors heme, arginine, BH(4) flavin adenine dinucleotide/flavin mononucleotide (FAD/FMN) and NADPH, via the far-reaching effects of the two Cbl coenzymes, methionine synthase (MS) and methylmalonyl CoA mutase (MCoAM) in, or on, the folate, glutathione, tricarboxylic acid (TCA) and urea cycles, oxidative phosphorylation, glycolysis and the pentose phosphate pathway. Deficiency of any of theNOS substrates and cofactors results in 'uncoupled' NOS reactions, decreasedNO production and increased or excessive O(2) (-), H(2)O(2), ONOO(-) and other reactive oxygen species (ROS), reactive nitric oxide species (RNIS) leading to pathology. (2) Cbl is also the overlooked ultimate determinant of positive glutathione status, which favours the formation of more benign NO species, s-nitrosothiols, the predominant form in which NO is safely deployed. Cbl status may consequently act as a 'back-up disc' that ensures the active status of antioxidant systems, as well as reversing and modulating the effects of nitrosylation in cell signal transduction.New evidence shows that GSCbl can significantly promote iNOS/ eNOS NO synthesis in the early stages of inflammation, thus lowering high levels of tumour necrosis factor-a that normally result in pathology, while existing evidence shows that in extreme nitrosative and oxidative stress, GSCbl can regenerate the activity of enzymes important for eventual resolution, such as glucose 6 phosphate dehydrogenase, which ensures NADPH supply, lactate dehydrogenase, and more; with human clinical case studies of OHCbl for cyanide poisoning, suggesting Cbl may regenerate aconitase and cytochrome c oxidase in the TCA cycle and oxidative phosphorylation. Thus, Cbl may simultaneously promote a strong inflammatory response and the means to resolve it.
    Journal of Nutritional & Environmental Medicine 02/2007; 16(3-4):181-211.
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    Carmen Wheatley
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    ABSTRACT: Cobalamin carrier proteins,the Transcobalamins (TCS), are elevated during trauma, infections and chronic inflammatory conditions. This remains un-explained. It is proposed that such TC elevations signal a need for cobalamin central to the resolution of inflammation. Thus Cobalamin may regulate the transcription factor, NFkappaB, activation or suppression of which determines the inflammatory response and its resolution. Such regulation may involve at least 5 separate mechanisms: (i) hormone-like regulation of TNFalpha, through reduction of excess NO by cobalamin, as well as through the selective inhibition, in tandem with glutathione, of inducible nitric oxide synthase; (ii) quenching of nitric oxide radicals and reactive oxygen species, enhanced by cobalamin's glutathione sparing effect; (iii) the promotion of acetylcholine synthesis, central to the neuro-immune cholinergic anti-inflammatory pathway; (iv) the promotion of oxidative phosphorylation; (v) and a bacteriostatic role of the TCS released by neutrophil secondary granules during phagocytosis, which also appears to modulate the inflammatory response. TC elevations are dependent on NFkappaB activation, through crosstalk between NFkappaB and Sp1, another member of the helix-loop-helix protein family, which directly mediates transcription of the TCII gene. Sp1 also has binding sites on the TNFalpha and EGF gene promoters. NFkappaB may thus ensure sufficient cobalamin to determine its own eventual suppression. Cobalamin's established regulation of EGF may additionally preserve normal function of macrophages and the coagulation cascade in wound healing. By regulating NFkappaB, Cobalamin may also be the as yet unidentified mediator needed to potentiate the anti-inflammatory action of eicosanoids derived from omega-3 essential fatty acids. Moreover, animal and human clinical data suggests that high dose cobalamin may prove a promising approach to SIRS/sepsis/septic and traumatic shock.
    Medical Hypotheses 02/2006; 67(1):124-42. · 1.18 Impact Factor
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    Carmen Wheatley
    01/2006;
  • Carmen Wheatley
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    ABSTRACT: NO ABSTRACT: Case History of an exceptional survival of untreated Multiple Myeloma Stage 1.
    01/2002: pages 267-312; , ISBN: 0-7432-0677-0
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    Carmen Wheatley
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    ABSTRACT: NO ABSTRACT. Introduction: “Multiple myeloma is incurable”, the textbooks tell us.(1, 2) Median survival depends on the stage at diagnosis and can range from six months to five years.(3) Long term survival is virtually unknown, but not quite. The 1992 survival curve for an unselected series of 156 patients at St Bartholomew's Hospital shows a 2.5% survival at 10 years.(4) A much earlier study at the Mayo Clinic, covering 870 cases, found a not dissimilar total survival rate at 10 years of 2.2%, with a 3% survival for those 597 patients diagnosed in the 1964 onwards, ...
    01/2002: pages 267-322; , ISBN: 0-7432-0677-0
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    Carmen Wheatley
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    ABSTRACT: NO ABSTRACT: Introduction The aetiology of Multiple Myeloma is generally acknowledged as obscure. (1) The epidemiology presents a number of puzzles. Most cancers are the affliction of age. Myeloma is more remarkably so, with few cases under the age of 40, (0.3% under 30), 98% above this demarcation line and a median incidence age of 65 years. (2) (Figure I). Whilst sporadic community(3) and familial(4,5) clusters have occurred, - and indeed, even husband and wife cases(6,7), - to date no evidence of genetic predisposition has been discovered(5), and failure to find a definitive environmental cause for the community clusters might suggest that such cases are simply random, given the statistical low incidence relative to Myeloma incidence in general.(3,7) Myeloma also affects more men than women, more blacks than whites, at a relatively earlier age than whites, and farmers more than the general population. (8) A French study noted a 40% excess prevalence, age- and sex-adjusted, amongst farmers, relative to other occupations.(9) Other at-risk groups include foresters(8), fishermen(8), veterinarians (10), teachers(11), anaesthesiologists(12), radiologists, and anyone exposed to ionising radiation.(8) Though the latter would seem an obvious risk factor, intriguingly the proportion of atom bomb victims at Hiroshima and Nagasaki who ultimately developed Myeloma seems relatively small, if still significant.(8) Indeed, Myeloma is a rare cancer, accounting for no more than 1% of all cancers, and 10% of all haematopoietic malignancies(2), although its incidence, as with the majority of all cancers, is on the increase.
    Journal of Orthomolecular Medicine 01/2002; 17(1).
  • Carmen Wheatley
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    ABSTRACT: NO ABSTRACT> Introduction:The etiology of Multiple Myeloma is generally acknowledged as obscure [1]. The epidemiology presents a number of puzzles. Most cancers are the affliction of age. Myeloma is more remarkably so, with few cases under the age of 40, (0.3% under 30), 98% above this demarcation line and a median incidence age of 65 years[2 ]. Whilst sporadic community [3] and familial [4,5] clusters have occurred, and indeed, even husband and wife cases,[6,7] to date no evidence of genetic predisposition has been discovered [5]. Failure to find a definitive environmental cause for the community clusters might suggest that such cases are simply random, given their statistical low incidence relative to Myeloma incidence in general [3,7]. Myeloma also affects more men than women, more blacks than whites and at a relatively earlier age than whites, farmers more than the general population [8]. A French study noted a 40% excess prevalence, age- and sex-adjusted, amongst farmers, relative to other occupations [9]. Other at-risk groups include foresters [8,] fishermen [8], veterinarians[10], teachers[11], anaesthesiologists [12], radiologists, and anyone exposed to ionizing radiation [8]. Though the latter would seem an obvious risk factor, intriguingly the pro- portion of atom bomb victims at Hiroshima and Nagasaki who ultimately developed Myeloma seems relatively small, if still significant [8]. Indeed, Myeloma is a rare cancer, accounting for no more than 1% of all cancers, and 10% of all haematopoietic malignancies [2], although its incidence, as with the majority of all cancers, is on the increase. It now appears that Multiple Myeloma, though a B cell malignancy, very much conforms to what we know about the mo- lecular biology of solid tumours, even down to the development of angiogenesis, albeit in the marrow [13]. The oncogenesis of Myeloma takes place over several decades in the largely silent, pre-cancer condition known as Monoclonal Gammopathy of Unknown Significance (MGUS), where the clonal plasma cells are immortalized but not transformed. About 24% of people with MGUS go on to develop full blown Multi- ple Myeloma and/or other lympho-prolif- erative malignancy [14]. MGUS and pre-Stage I Myeloma are relatively indolent, benevolent phases. Paradoxically, however, once in the active phase, Myeloma is lethal and without a known cure, with the exception of allogeneic transplants which carry a 41% mortality rate [15]. Moreover, paradoxes and problems abound in the pathology and treatment of Myeloma, a mystifyingly heterogenous disease, with survivals recorded ranging from a few months to nearly two decades [2]. Con- ventional chemotherapy offers a median survival of 18 months to 2 years. Newer approaches of high-dose combination chemotherapy and autologous stem cell transplant have demonstrated an improved median survival of 3 to 4 years, with 20% alive at 5 years. Improvement is pertinent largely to a minority of patients who fit particular diagnostic criteria, including age less than 56 years, low ß2 microglobulin, low C-Reactive Protein, no deletion of chromosome 13 and a low Labeling Index [16]. Myeloma, in fact, presents a therapeutic enigma. It does not respond to therapy as cancers generally do [17]. It does not exhibit a dose-response effect; remission duration and survival does not appear directly re- lated to Myeloma cell-kill; maintenance therapy does not necessarily prolong remission and survival duration; surviving Myeloma cells do not necessarily begin to grow and proliferate exponentially when treatment stops; and treatment can lower the M-protein to a plateau beyond which it will not fall lower despite continued therapy. Finally, in the rare cases where longterm survival is achieved beyond ten years, there is also the paradox that, alone of all hae- matopoietic malignancies, such longterm survival does not equal cure, and relapse is still, bafflingly, the rule [18]. Why? Why too, for instance, should Myeloma show “a special predilection for the spinal column”?[19] Again, the mysterious, rare but recognized phenomenon of the plateau achieved without treatment, peculiar to Myeloma, in which the disease is still present but “spontaneously” becomes inactive [20], suggests an inner mechanism of control. If we knew what this mechanism involved, we might be able to access it and perhaps prolong the plateau indefinitely, as good as a cure. Similarly, in a tiny minority, about 4%,20 MGUS occasionally disappears altogether. If we knew why, as Robert Kyle has remarked, “If we could reverse MGUS, we could cure Myeloma.” [21].
    Journal of Orthomolecular Medicine 01/2002; Vol.17(1-0834-4825):7-16.
  • Carmen Wheatley
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    ABSTRACT: The adverse and contradictory results of recent intervention trials are little understood. The reason for this may be that the trials were designed on mistaken premises. The drugs-testing paradigm was adopted, and no consideration was given to the nature of nutrients. Nutrients do not act in isolation, unlike drugs. They are dependent on synthetic action for their proper effects. This can lead to paradoxes, knowledge of which must be incorporated in the design of a good intervention trial. Dose, isomeric form, diet, nutrient biochemistry, genetic population difference and the use of exact bio-markers, are all key considerations in the design of a good trial. More precise screening for effective chemo-preventative and therapeutic agents is urged and a new 'nutrient paradigm' trial protocol, taken from the new field of 'nutraceuticals', with a relatively active placebo arm, is proposed. Read More: http://informahealthcare.com/doi/abs/10.1080/13590849862069
    Journal of Nutritional & Environmental Medicin. 07/1998; 8(3-1364-690):277-288.
  • Carmen Wheatley
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    ABSTRACT: The adverse and contradictory results of recent intervention trials are little understood. The reason for this may be that the trials were designed on mistaken premises. The drugs-testing paradigm was adopted, and no consideration was given to the nature of nutrients. Nutrients do not act in isolation, unlike drugs. They are dependent on synthetic action for their proper effects. This can lead to paradoxes, knowledge of which must be incorporated in the design of a good intervention trial. Dose, isomeric form, diet, nutrient biochemistry, genetic population difference and the use of exact bio-markers, are all key considerations in the design of a good trial. More precise screening for effective chemo-preventative and therapeutic agents is urged and a new 'nutrient paradigm' trial protocol, taken from the new field of 'nutraceuticals', with a relatively active placebo arm, is proposed.
    Journal of Nutritional & Environmental Medicine 07/1998; 8(3-IND21972445):277-288.
  • C Wheatley
    The Lancet 07/1997; 349(9068):1844-5. · 39.06 Impact Factor
  • Carmen Wheatley
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    ABSTRACT: Results from intervention trials of β-carotene, α-tocopherol, and vitamin A (eg, CARET and ATBC 1) have brought much adverse publicity for vitamins and nutritional supplements. Parties with a vested interest have jumped on board, clamouring for bans and legislation. But no one has questioned the fundamental premise of these trials—namely, that vitamins are being tested and treated as if they were drugs. This paradigm is misguided and largely accounts for the conclusions of the recent trials.
    The Lancet 06/1997; 349(9068):1844. · 39.06 Impact Factor

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